This application addresses: Broad Challenge Area: (15) Translational Science Topic: 15-OD (ORDR)-101* Pilot projects for prevention, early detection and treatment of rare diseases. Exon capture and large-scale sequencing for disease-cause identification, early detection and drug discovery in nephrotic syndrome. Background: Chronic kidney diseases (CKD) take one of the highest tolls on human health. They insidiously lead to end- stage kidney disease (ESKD) requiring dialysis or kidney transplantation for survival. 20 million individuals in the U.S. suffer from CKD with a treatment cost of >$20 billion/yr. Steroid-resistant nephrotic syndrome (SRNS) is a rare disease that constitutes the second most frequent cause of ESKD in children and young adults. No curative treatment is available. We demonstrated that many pediatric cases with SRNS are rare recessive single-gene disorders. 25% of all SRNS cases are caused by recessive NPHS2/podocin mutations and many additional single-gene causes of SRNS exist. Identification of rare single-gene causes for SRNS has provided fundamental insights into disease mechanisms of nephrotic syndrome in children and adults. It has allowed to perform unequivocal molecular genetic diagnostics for early detection of SRNS (www.renalgenes.org) and to stratify patient cohorts for therapeutic trials. The study of recessive disease mechanisms is particularly powerful, as they can be recapitulated in animal models by gene knockdown/knockout in zebrafish or mice. These animal models allow for high throughput drug screening to define new molecular targets for treatment. Knowledge Gap and New Technology: The current knowledge gap in disease causes of SRNS has hampered diagnostics, early detection and target- oriented drug treatment for this incurable disease. The heterogeneity of recessive SRNS-causing genes, and their rarity, pose severe limitations to gene identification. Recently, we introduced a new technology into gene discovery of rare recessive single-gene causes by developing a combined approach of homozygosity mapping (HM) with consecutive exon capture and large-scale sequencing. Feasibility and Existing Resources: We have ascertained DNA samples and clinical data from over 2,000 families with SRNS world-wide and have clarified the molecular cause of SRNS in 15% of cases. We applied HM successfully to the identification of 11 novel CKD genes. Recently, we demonstrated that HM is broadly applicable even to single cases with rare diseases. In 30-80% of cases it yielded homozygous candidate regions, which contained the disease causing homozygous mutation in 93% of cases. When identifying by HM recessive PLCE1 mutations as a rare cause of SRNS (~1%), we recapitulated the disease in zebrafish, thereby generating a model for drug screening. By total genome HM in 250 of the 2,000 families with SRNS (using 250k and 1Mill SNP arrays) we yielded new homozygous candidate regions in 100/250 families (40%), and demonstrated that more than 20 additional unknown recessive single-gene causes of SRNS must exist. Thus, we will address the following specific aims: Specific Aims SA1. Identify novel causes of the rare disease steroid-resistant nephrotic syndrome (SRNS) by total genome homozygosity mapping, with consecutive targeted exon capture and large-scale sequencing. SA2. Establish zebrafish models of novel SRNS genes and test new treatment approaches. SA3. Rapidly expand the new strategy of gene identification to other autosomal recessive conditions with a urogenital phenotype. Significance: The new approach that we developed will have the following impact for SRNS and other rare diseases: i) Gene identification will generate novel insights into the etiology and disease mechanisms of SRNS. ii) It will allow screening and early detection for this rare disease and etiologic stratification for clinical trials. iii) Gene identification will generate novel molecular targets to treat SRNS, which currently is incurable. iv) Further development of disease models in zebrafish for SRNS-causing genes will enable high throughput screening assays for drug discovery of novel compounds to treat SRNS. We already demonstrated the feasibility of this approach. The project will expand our successful preliminary results of HM with consecutive targeted exon capture and large-scale sequencing to a large number of families with rare diseases. Following this ARRA, we will enlarge the project (with existing resources) to study all recessive pediatric diseases at the U of M Mott Children's Hospital and beyond. These studies, for which all resources and technologies are in place, would greatly benefit from an influx of funds to quickly and significantly advance disease-cause identification in SRNS and other rare diseases by rapidly generating data using highly efficient, but costly, new technology of exon capture and large-scale sequencing, which we have established. Chronic kidney diseases take one of the highest tolls on human health. Steroid-resistant nephrotic syndrome (SRNS) is a rare disease that constitutes the second most frequent cause of ESKD in children and young adults. No curative treatment is available. 25% of all SRNS cases are caused by recessive NPHS2/podocin mutations, and we recently demonstrated, by genetic mapping, that many additional single-gene causes of SRNS must exist. Identification of rare single-gene causes for SRNS has provided fundamental insights into disease mechanisms of nephrotic syndrome in children and adults. Recently, we introduced a new technology into gene discovery of rare recessive single-gene causes by developing a combined approach of homozygosity mapping with consecutive exon capture and large-scale sequencing. We have ascertained DNA samples and clinical data from over 2,000 families with SRNS world- wide and have clarified the molecular cause of SRNS in 15% of cases. We here propose to 1) Identify novel causes of the rare disease SRNS by total genome homozygosity mapping, with consecutive targeted exon capture and large-scale sequencing;2) Establish zebrafish models of novel SRNS genes and test novel treatment approaches;and 3) Rapidly expand the new strategy of gene identification to other autosomal recessive conditions with a urogenital phenotype.